Pulse Radiolysis Studies of Temperature Dependent Electron Transfers among Redox Centers in ba 3 -Cytochrome c Oxidase from Thermus thermophilus : Comparison of A- and B-Type Enzymes.

The functioning of cytochrome oxidases involves orchestration of long-range electron transfer (ET) events among the four redox active metal centers. We report the temperature dependence of electron transfer from the Cu site to the low-spin heme-() site, i.e., Cu + heme-() → Cu + heme-() in three structurally characterized enzymes: A-type from (PDB code 3HB3) and bovine heart tissue (PDB code 2ZXW), and the B-type from (PDB codes 1EHK and 1XME). , data sets were obtained with the use of pulse radiolysis as described previously. Semiclassical Marcus theory revealed that λ varies from 0.74 to 1.1 eV, , varies from ∼2 × 10 eV (0.16 cm) to ∼24 × 10 eV (1.9 cm), and β varies from 9.3 to 13.9. These parameters are consistent with diabatic electron tunneling. The II-Asp111Asn Cu mutation in cytochrome had no effect on the rate of this reaction whereas the II-Met160Leu Cu-mutation was slower by an amount corresponding to a decreased driving force of ∼0.06 eV. The structures support the presence of a common, electron-conducting "wire" between Cu and heme-(). The transfer of an electron from the low-spin heme to the high-spin heme, i.e., heme-() + heme- → heme-() + heme-, was not observed with the A-type enzymes in our experiments but was observed with the ; its Marcus parameters are λ = 1.5 eV, = 26.6 × 10 eV (2.14 cm), and β = 9.35, consistent also with diabatic electron tunneling between the two hemes. The II-Glu15Ala mutation of the K-channel structure, ∼ 24 Å between its CA and Fe-, was found to completely block heme- to heme- electron transfer. A structural mechanism is suggested to explain these observations.